Temporal and spatial expression of <i>CYP15C1</i>.

<p>(A) qRT-PCR analysis of the spatial expression of <i>CYP15C1</i> in the silkworm strain Kinshu×Showa. “<i>CYP15C1</i>/<i>rp49</i>” on the vertical axis indicates the level of <i>CYP15C1</i> mRNA normalized to that of internal <i>rp49</i> mRNA. RNAs were collected from larvae on day 1 of the fourth instar (4th D1), fourth instar larvae showing head capsule slippage (4th HCS), larvae on day 2 of the fifth instar (5th D2), and larvae on day 1 after the onset of spinning (Spin+1). CC-CA, corpus cardiacum-corpus allatum complex; PG, prothoracic gland; Br, brain; FB, fat body; MG, midgut; Ep, epidermis; Ms, muscle; Mp, Malpighian tubule; SiG, silk gland; SaG, salivary gland; Ts, testis; and Ov, ovary. (B) <i>In situ</i> mRNA hybridization of <i>CYP15C1</i> and <i>JHAMT</i>. Br-CC-CA complexes on day 2 of the fourth instar and day 4 of the fifth instar were used for analysis. Signals of both genes were limited to CA as indicated by arrows, but <i>JHAMT</i> was not detected on day 4 of the fifth instar. The purple coloration in the brain is primarily due to ommochrome pigments and does not reflect gene expression. The result of control experiments using sense probes are shown in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002486#pgen.1002486.s002" target="_blank">Figure S2</a>. (C) Developmental changes in the rate of JH biosynthesis by <i>B. mori</i> CA <i>in vitro</i>. The data are based on Kinjoh et al. (2007). Black, red, and blue lines indicate CA from unsexed larvae, female and male animals, respectively. The activity in CA on day 1 of the fourth instar was set as 100. (D) Temporal expression patterns of <i>JHAMT</i> (upper) and <i>CYP15C1</i> (lower) in the Br-CC-CA (first and second instar larvae) or CC-CA (third to fifth instar larvae, pupae, and adults) complex. Developmental stages are defined as h/days after certain developmental events [i.e., molting, head capsule slippage (HCS), spinning, or emergence] or by a spiracle index (si) <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002486#pgen.1002486-Kiguchi1" target="_blank">[56]</a>. Animals were unsexed during larval stages, while sexed during pupal and adult stages (female in red and male in blue). The expression profile of <i>JHAMT</i> after the second larval instar is based on published data (20). Expression levels measured on day 2 of the 4th larval instar are arbitrarily set at 100 (for actual transcript numbers per <i>rp49</i>) and are shown in a log scale. Asterisks indicate that data were not available.